Angiogenesis and osteogenesis are tightly coupled during bone development and regeneration. Mesenchymal cells in the developing stroma elicit angiogenic signals to attract new blood vessels to bone rudiments or regenerates. Other signals, likely emanating from the incoming vasculature, stimulate bone cell specification through interactions with cells within the vascular stem cell niche. The long-term goal of this project is to understand the cellular and molecular mechanisms that control interactions between bone cells and vascular elements. During the past funding period, we have identified the hypoxia inducible factor alpha (HIF-11) pathway as a key component in this process. Using a genetic approach, we have demonstrated that overexpression of HIF1 in mouse osteoblasts through disruption of Vhl profoundly increases angiogenesis and osteogenesis; these processes appear to be coupled by cell non-autonomous mechanisms involving VEGF. Preliminary studies described below suggest that endothelial cells are one source of bone morphogenic signal which couples angiogenesis to bone formation. Angiogenesis also appears to drive osteogenesis when long bones from the mutant and wild type mice are subjected to injury (distraction osteogenesis). Surprisingly however, manipulation of HIF1 levels in mature osteoblasts does not influence the formation of the flat bones of the skull. These results suggest that the mechanisms which couple angiogenesis to osteogenesis are context (skeletal site) specific. In this renewal application, we propose studies to characterize cellular and molecular mechanisms responsible for coupling angiogenesis to osteogenesis in three different skeletal contexts. This renewal grant has three aims: Specific Aim 1: Angiogenic-osteogenic coupling in developing long bones. Specific Aim 2: Angiogenic-osteogenic coupling following bone injury. Specific Aim 3: Angiogenic-osteogenic coupling in the developing cranium. The ultimate goal of this project is to elucidate the functional mechanisms underlying HIF-1/VEGF stimulated angiogenesis and osteogenesis. These findings should lead to better understanding of precise communication between angiogenesis and osteogenesis and aid in the design of new therapies to accelerate bone healing following injury